Recognition of microbial antigens by the host immune system is mediated through innate immune receptors, whose activation represents an important step in the initiation of an inflammatory response. Toll-Like Receptors (TLR) are a family of innate immune receptors that play a crucial role in mediating an immune response to foreign antigens. TLR3s are pathogen associated molecular pattern recognition receptors that recognize double-stranded RNA (dsRNA) as well as the synthetic dsRNA analog poly-riboinosinic-ribocytidylic acid (poly(I:C)). See e.g. Alexopoulou et al., 413 Nature 732 (2001)). Moreover, TLR3 has been shown to recognize endogenous ligands such as mRNA released from necrotic cells suggesting that necrotic cell death at inflammation sites may contribute to activation of TLR3. See e.g. Kariko et al., 26 J. Biol. Chem. 12542 (2004).
Activation of TLR3s by dsRNA or poly(I:C) ligands induces secretion of pro-inflammatory cytokines and chemokines and can modulate disease outcomes during infection-associated inflammation. Importantly, TLR3 activation in vivo occurs in the context of viral infection or necrosis associated with inflammation. See Tabeta et al., 101 Proc. Natl. Acad. Sci. USA 3516 (2004)); see also Kariko et al., 26 J. Biol. Chem. 12542 (2004). For example, the human TLR3 peptide chain is expressed in the central nervous system (CNS), where it is required to control infection by the HSV-1 virus, which spreads from the epithelium to the central nervous system via cranial nerves to cause herpes simplex encephalitis in TLR3 deficient patients. See e.g. Zhang et al., 317 Science 1522 (2007). Furthermore, human TLR3 peptide chain activation results in inflammatory responses associated with pathological conditions such as, for example, primary biliary cirrhosis of liver tissues. See e.g. Takii et al., 85 Lab. Invest. 836 (2005).
Overall, these data demonstrate that activation of TLR3 initiates cascades of phosphorylation and transcriptional activation events that result in the production of numerous inflammatory cytokines that contribute to innate immunity (reviewed by Takeda and Akira, J. Derm. Sci. 34:73-82 (2004)). Further, these data indicate that sustained TLR3 activation is a critical component in the modulation of infection-associated inflammatory diseases. Published data lend further support to this hypothesis as shown by findings that associate over-production of pro-inflammatory cytokines to systemic inflammatory response syndrome, infection-associated acute cytokine storms (reviewed by Van Amersfoort et al., Clin. Microbiol. Rev. 16: 379-414 (2003)) and immune-mediated chronic conditions such as rheumatoid arthritis (reviewed by Miossec et al., Curr. Opin. Rheumatol. 16:218-222 (2004)) and inflammatory bowel diseases (reviewed by Ogata and Hibi, Curr. Pharm. Des. 9: 1107-1113 (2003)).
Currently, a number of different therapeutic approaches have been taken to target the activity of TLR3s for treatment of different indications. These TLR3 therapeutics include human peptide chain based TLR3 therapeutics, monoclonal antibody antagonists of TLR3, and TLR3 ligand agonists such as dsRNA, poly(I:C) as well as functional analogs of these that target TLR3 activity. The potential indications for monoclonal antibody antagonist based TLR3 therapeutics include inflammatory conditions, sepsis, inflammatory bowel disease, inflammatory pulmonary disease, and autoimmune diseases. The potential indications and uses for TLR3 therapeutics that are agonists include post-viral fatigue syndrome, glioma, prostate cancer, antiviral vaccines, bladder cancer, cervical dysplasia, human papilloma virus infection, breast cancer, viral infection prevention, tissue regeneration, and avian influenza vaccines.
Extensive safety testing will be required before any TLR3 therapeutic for human use can be brought to the market place. Such safety testing will involve both in vivo safety testing in animal models as well as the in vitro testing of TLR3 therapeutics. For example, antibody based TLR3 therapeutics may require the generation of surrogate antibodies against a TLR3 peptide chain expressed by a particular model animal as well as significant in vitro characterization of such surrogate antibodies. Such surrogate generation and in vitro characterization will require the use of TLR3 polynucleotides and peptide chains from a suitable model animal. Importantly, the identification of suitable animal models for such safety testing requires the identification of animal species capable of expressing a TLR3 with high identity and homology to human TLR3 (SEQ ID NO: 13).
Thus, a need exists for the identification of polynucleotides encoding TLR3s and TLR3 peptide chains capable of being expressed in an animal model suitable for the safety testing of TLR3 therapeutics. A need also exists for related methods such as methods of expressing peptide chains and testing the safety of a TLR3 therapeutic in an animal model identified as suitable for safety assessment of TLR3 therapeutics.